Quantitative Colocalization Analysis of Confocal Fluorescence Microscopy Images

Colocalization is an important finding in many cell biological studies. This unit describes a protocol for quantitative evaluation of images with colocalization based on the calculation of a number of specialized coefficients. First, images of double‐stained sections are subjected to background correction. Then, various coefficients are calculated. Meanings of the coefficients and a guide to interpretation of their results indicating either presence or absence of colocalization are given. Success in colocalization studies depends on the quality of analyzed images, proper preparation of the images for coefficients calculations, and correct interpretation of obtained results. This protocol helps to ensure reliability of colocalization coefficients calculations. Curr. Protoc. Cell Biol. 52:4.16.1‐4.16.16. © 2011 by John Wiley & Sons, Inc.

[1]  H. Favoreel,et al.  Porcine Sialoadhesin (CD169/Siglec-1) Is an Endocytic Receptor that Allows Targeted Delivery of Toxins and Antigens to Macrophages , 2011, PloS one.

[2]  Vadim Zinchuk,et al.  Recent advances in quantitative colocalization analysis: focus on neuroscience. , 2009, Progress in histochemistry and cytochemistry.

[3]  Tokiko Watanabe,et al.  Involvement of host cellular multivesicular body functions in hepatitis B virus budding , 2007, Proceedings of the National Academy of Sciences.

[4]  M. Steer,et al.  Cause-effect relationships between zymogen activation and other early events in secretagogue-induced acute pancreatitis. , 2007, American Journal of Physiology - Gastrointestinal and Liver Physiology.

[5]  D. Deforce,et al.  Tyrosine phosphorylation and lipid raft association of pseudorabies virus glycoprotein E during antibody-mediated capping. , 2007, Virology.

[6]  S. Krisans,et al.  IDI2, a Second Isopentenyl Diphosphate Isomerase in Mammals* , 2007, Journal of Biological Chemistry.

[7]  S. Milam,et al.  Integrins regulate opioid receptor signaling in trigeminal ganglion neurons , 2007, Neuroscience.

[8]  G Ulrich Nienhaus,et al.  Affinity of C-reactive protein toward FcgammaRI is strongly enhanced by the gamma-chain. , 2007, The American journal of pathology.

[9]  R. Sanyal,et al.  Activation-induced endocytosis of the raft-associated transmembrane adaptor protein LAB/NTAL in B lymphocytes: evidence for a role in internalization of the B cell receptor. , 2006, International immunology.

[10]  S. H. Young,et al.  Requirement of the TRPC1 Cation Channel in the Generation of Transient Ca2+ Oscillations by the Calcium-sensing Receptor* , 2006, Journal of Biological Chemistry.

[11]  P. Insel,et al.  Microtubules and Actin Microfilaments Regulate Lipid Raft/Caveolae Localization of Adenylyl Cyclase Signaling Components* , 2006, Journal of Biological Chemistry.

[12]  H. Patel,et al.  Role of 12-lipoxygenase in volatile anesthetic-induced delayed preconditioning in mice. , 2006, American journal of physiology. Heart and circulatory physiology.

[13]  P. Insel,et al.  Protection of adult rat cardiac myocytes from ischemic cell death: role of caveolar microdomains and delta-opioid receptors. , 2006, American journal of physiology. Heart and circulatory physiology.

[14]  P. Insel,et al.  Focal Adhesions in (Myo)fibroblasts Scaffold Adenylyl Cyclase with Phosphorylated Caveolin* , 2006, Journal of Biological Chemistry.

[15]  G. Gerken,et al.  Trypsin-Sensitive Modulation of Intestinal Epithelial MD-2 as Mechanism of Lipopolysaccharide Tolerance1 , 2006, The Journal of Immunology.

[16]  F. Dombrowski,et al.  Tauroursodeoxycholic acid inserts the bile salt export pump into canalicular membranes of cholestatic rat liver , 2006, Laboratory Investigation.

[17]  A. J. North,et al.  Seeing is believing? A beginners' guide to practical pitfalls in image acquisition , 2006, The Journal of cell biology.

[18]  M. Beckerle,et al.  Atrial natriuretic peptide promotes cardiomyocyte survival by cGMP-dependent nuclear accumulation of zyxin and Akt. , 2005, The Journal of clinical investigation.

[19]  T. Okada,et al.  Experimental LPS‐induced cholestasis alters subcellular distribution and affects colocalization of Mrp2 and Bsep proteins: A quantitative colocalization study , 2005, Microscopy research and technique.

[20]  M. Nguyen,et al.  Tumor metastasis but not tumor growth is dependent on Src-mediated vascular permeability. , 2005, Blood.

[21]  V. Zinchuk,et al.  Direct action of platelet activating factor (PAF) induces eosinophil accumulation and enhances expression of PAF receptors in conjunctivitis. , 2005, Molecular vision.

[22]  A. Fukushima,et al.  Dynamics of PAF-induced conjunctivitis reveals differential expression of PAF receptor by macrophages and eosinophils in the rat , 2004, Cell and Tissue Research.

[23]  Lukas Landmann,et al.  Colocalization analysis yields superior results after image restoration , 2004, Microscopy research and technique.

[24]  H. Seguchi,et al.  Asynchronous expression and colocalization of Bsep and Mrp2 during development of rat liver. , 2002, American journal of physiology. Gastrointestinal and liver physiology.

[25]  A. Smallcombe,et al.  Multicolor imaging: the important question of co-localization. , 2001, BioTechniques.

[26]  J. Lippincott-Schwartz,et al.  Studying protein dynamics in living cells , 2001, Nature Reviews Molecular Cell Biology.

[27]  D. Keppler,et al.  Conjugate export pumps of the multidrug resistance protein (MRP) family: localization, substrate specificity, and MRP2-mediated drug resistance. , 1999, Biochimica et biophysica acta.

[28]  P. Meier,et al.  The Sister of P-glycoprotein Represents the Canalicular Bile Salt Export Pump of Mammalian Liver* , 1998, The Journal of Biological Chemistry.

[29]  J. Aten,et al.  Measurement of co‐localization of objects in dual‐colour confocal images , 1993, Journal of microscopy.

[30]  Elmer S. West.  From the U. S. A. , 1965 .